Resurfaced Shape Complementary Proteins That Selectively Bind the Oncoprotein Gankyrin

[Image: see text] Increased cellular levels of protein–protein interactions involving the ankyrin repeat oncoprotein gankyrin are directly linked to aberrant cellular events and numerous cancers. Inhibition of these protein–protein interactions is thus an attractive therapeutic strategy. However, the relatively featureless topology of gankyrin’s putative binding face and large surface areas involved in gankyrin-dependent protein–protein interactions present a dramatic challenge to small molecule discovery. The size, high folding energies, and well-defined surfaces present in many proteins overcome some of the challenges faced by small molecule discovery. We used split-superpositive Green Fluorescent Protein (split-spGFP) reassembly to screen a 5 × 10(9) library of resurfaced proteins that are shape complementary to the putative binding face of gankyrin and identified mutants that potently and selectively bind this oncoprotein in vitro and in living cells. Collectively, our findings represent the first synthetic proteins that bind gankyrin and may represent a general strategy for developing protein basic research tools and drug leads that bind disease-relevant ankyrin repeats.